Tuesday, March 5, 2013

I recently had the pleasure of reading about the succession of microbes in flowers as they emerge, open, and eventually fall from the tree. This interesting paper (published by the Handelsman group and found here:http://mbio.asm.org/content/4/2/e00602-12) used 454 pyrosequencing to identify bacterial community members in flowers across 5 different trees throughout time. I especially liked figure 4 (below):

They used the appearance of different groups at different parts of the season to identify "poineer", early, mid, late, climax and "generalist" microbes. Although the list of sequences or microbes in each of these groups isn't provided, they go on to state: “Late” group members peaked when flowers had been open for 3 days and included a high abundance of Lactobacillusand Acetobacter taxa, whose occurrences aligned with previously reported conditions of flower decomposition by yeast" I was surprised that beyond a casual mention in the paper's discussion, there was no attempt to probe whether or not the communities found on flowers resembled those found associated with pollinators.

As it turns out, the genera Lactobacillus and Acetobacter are also associated with sweat bees, honey bees and bumble bees (see http://aem.asm.org/content/early/2013/01/03/AEM.03681-12.abstract and http://www.biomedcentral.com/1471-2180/12/221). I downloaded the entire apple flower dataset from MG-RAST and checked to see if any would affiliate with the bee specific sequences (using a 60% threshold). Here is a quick table below, sorted alphabetically by "family" level. Out of a total of ~35,000 sequences, a subset (~5,600) have been previously found in bees.

Family

Count

Acetobacetraceae

9

Acetobacteraceae

663

Actinomycetales

713

Actinomycetales_incertae_sedis

1271

alpha-1

53

alpha-2.2

11

alpha-2.2

47

Anaplasmataceae

1

beta

6

Bifidobacteriaceae

54

Brucellaceae

288

Burkholderiaceae

66

Burkholderiales_bacterium_YT0099

92

Comamonadaceae

38

Comamonadaceae_incertae_sedis

19

Comamonodaceae_incertae_sedis

194

Enterobacteriaceae

636

firm-4

3

gamma-1

3

Lactobacillaceae

293

Moraxellaceae

55

Neisseriaceae

19

Oxalobacteraceae

520

Staphylococcaceae

560

This picture changes when you classify the dataset using a more "complete" training set - the SILVA+Bees training set we developed (http://www.biomedcentral.com/1471-2180/12/221). Sequences that found homes in the honey bee specific training set alone (within the honey bee specific clades), now find homes elsewhere and very few remain (highlighted below in red). It's also worth mentioning that one major conclusion of the paper -- the "novel" sequences uncovered by the experiment -- was true in my hands as well. A very large fraction of the sequences (~16,000) were "unclassified" at the family level, even at a 60% threshold using the NBC-RDPII and the SILVA + honey bee specific training set (see classifications below).

Acetobacteraceae

821

Acholeplasmataceae

1

Acidimicrobiaceae

1

Acidothermaceae

5

Actinomycetaceae

16

Actinomycetales_incertae_sedis

1

Actinospicaceae

1

Actinosynnemataceae

1

Aerococcaceae

18

Aeromonadaceae

5

Alcaligenaceae

55

Anaplasmataceae

3

Aurantimonadaceae

15

Bacillaceae

344

Bacillales_incertae_sedis

7

Bacteriovoracaceae

29

Bdellovibrionaceae

19

Beijerinckiaceae

28

beta

4

Bifidobacteriaceae

1

Bradyrhizobiaceae

118

Brevibacteriaceae

6

Brucellaceae

8

Burkholderiaceae

89

Burkholderiales_bacterium_YT0099

3

Burkholderiales_incertae_sedis

91

Caldilineaceae

2

Campylobacteraceae

5

Carnobacteriaceae

13

Caryophanaceae

1

Caulobacteraceae

322

Cellulomonadaceae

90

Chitinophagaceae

1357

Clostridiaceae

113

Comamonadaceae

264

Conexibacteraceae

112

Coriobacteriaceae

10

Corynebacteriaceae

79

Cryomorphaceae

8

Cyclobacteriaceae

127

Cystobacteraceae

17

Cytophagaceae

1357

Deinococcaceae

696

Dermabacteraceae

3

Dermatophilaceae

32

Desulfovibrionaceae

1

Dietziaceae

21

Enterobacteriaceae

570

Enterococcaceae

24

Erysipelotrichaceae

69

Erythrobacteraceae

288

Eubacteriaceae

6

Flammeovirgaceae

12

Flavobacteriaceae

795

Fusobacteriaceae

2

gamma-1

1

Gemmatimonadaceae

236

Geobacteraceae

9

Geodermatophilaceae

75

Glycomycetaceae

1

Gracilibacteraceae

6

Haliangiaceae

1

Holophagaceae

4

Hydrogenophilaceae

3

Hyphomicrobiaceae

138

Hyphomonadaceae

13

Iamiaceae

161

Ilumatobacter

19

Incertae_Sedis_XI

33

Incertae_Sedis_XII

3

Incertae_Sedis_XIII

3

Incertae_Sedis_XIV

7

Incertae_Sedis_XVIII

2

Intrasporangiaceae

386

Kineosporiaceae

44

Lachnospiraceae

60

Lactobacillaceae

422

Leptotrichiaceae

5

Leuconostocaceae

21

Methylobacteriaceae

188

Methylocystaceae

3

Methylophilaceae

12

Microbacteriaceae

340

Micrococcaceae

178

Micrococcineae_incertae_sedis

5

Micromonosporaceae

23

Moraxellaceae

190

Mycobacteriaceae

31

Myxococcaceae

9

Nakamurellaceae

23

Nannocystaceae

18

Neisseriaceae

15

Nitriliruptoraceae

2

Nitrosomonadaceae

27

Nitrospiraceae

15

Nocardiaceae

45

Nocardioidaceae

406

Nocardiopsaceae

11

Oxalobacteraceae

396

Paenibacillaceae

90

Parachlamydiaceae

9

Pasteurellaceae

4

Peptococcaceae

11

Peptostreptococcaceae

117

Phyllobacteriaceae

110

Planococcaceae

162

Polyangiaceae

38

Promicromonosporaceae

1

Propionibacteriaceae

39

Pseudomonadaceae

21

Pseudonocardiaceae

92

Rhizobiaceae

42

Rhodobacteraceae

350

Rhodobiaceae

3

Rhodocyclaceae

38

Rhodospirillaceae

57

Rhodothermaceae

30

Rickettsiaceae

30

Rubrobacteraceae

4

Ruminococcaceae

35

Sanguibacteraceae

57

Saprospiraceae

17

Sinobacteraceae

86

Sneathiellaceae

1

Solirubrobacteraceae

44

Sphaerobacteraceae

10

Sphingobacteriaceae

401

Sphingomonadaceae

1673

Spirochaetaceae

2

Sporichthyaceae

4

Staphylococcaceae

19

Streptococcaceae

42

Streptomycetaceae

26

Streptophyta

28

Streptosporangiaceae

3

Syntrophobacteraceae

1

Syntrophomonadaceae

2

Thermoactinomycetaceae

77

Thermoanaerobacteraceae

7

Thermomonosporaceae

5

Trueperaceae

2107

Is this a surprising result? If pollinator-affiliated sequences were found in abundance on the flowers you could explain it by the simple fact that flowers are visited by these insects and therefore bacteria from bees might be shed onto the flower parts. The relative paucity suggests instead that these bacteria, although inoculated into the environment by the bees, are specific to their hosts and perhaps do not thrive on the flower.